Paul ion trap mass spectrometers comprise a hyperbolic ring electrode and two rotationally symmetric hyperbolic end cap electrodes. If an electric voltage is applied to the end caps, on the one hand, and to the ring electrode, on the other, an essentially quadrupole field is generated in the interior. If the voltage is an RF voltage, then the RF electric field created is able to store ions. For practical reasons, it is usually the case that this RF storage voltage is only applied to the ring electrode, while the end cap electrodes are kept at ground potential. The RF storage voltage has a frequency which is usually around one megahertz.
According to Hans Dehmelt, the RF storage field can be envisaged as a pseudopotential well with a parabolic potential minimum in the center; the ions in the potential well are able to orbit on ellipses or oscillate through the center. The pseudopotential is a temporal integration over the square of the field intensity; the gradient of the pseudopotential continually drives the ions back to the center of the ion trap.
The ions are only stored when they have a mass above a cut-off mass, however. The term “mass” here is always to be understood as the charge-related mass m/z, as is required in mass spectrometry, i.e., the physical mass m divided by the number z of the (positive or negative) elementary charges. Ions below the cut-off mass are so light that during one half-phase of the RF storage voltage they can already be accelerated up to the opposite electrodes; temporal integration is no longer possible for them.
The remaining ions oscillate in the pseudopotential well in the ion trap, the oscillation frequencies being roughly inversely proportional to their mass. There are good approximation formulae for the relationship between mass and oscillation frequency. The oscillation frequencies are one characteristic for the mass; for example, the oscillations of the ions can be resonantly excited with very accurate mass selectivity.
If the ion trap is filled with a collision gas at a pressure between 101 and 103 Pascal, then the oscillations of the ions in the potential well are damped within a short time in such a way that the ions collect in a small cloud in the minimum of the potential well. The size of the cloud is determined by the Coulomb repulsion between the ions themselves, on the one hand, and by the centrally-directed force of the pseudopotential, on the other. The time required by the damping is inversely proportional to the pressure of the collision gas. At a pressure of around 102 Pascal, the time up to the damping is a few milliseconds; the ion undergoes a few hundred collisions in this time.
To measure fragment ion spectra in ion trap mass spectrometers, it is necessary to first select an ion species which one wishes to fragment into fragment ions and then measure. The fragment ions (of the first generation of fragmentations) are frequently termed “daughter ions”, and the ion species to be selected for the fragmentation is frequently termed “parent ions”. After selecting the parent ions, all other ions located in the ion trap are ejected from it so that only the parent ions remain. The parent ions do not have to have precisely the same mass; they can also be the different ions which have the same molecular formula of the elemental composition but include all the various isotopic combinations.
The process of ejecting all ions not selected is frequently termed “isolation” of the parent ions. The basic principles of the ejection are largely known and can easily be conducted in all commercially available ion trap mass spectrometers. It is based, on the one hand, on using the lower mass limit to eject the ions that are lighter than the parent ions and, on the other, using a mass-selective resonant excitation of the oscillations of the undesired heavier ions; the excitation process used is so strong that the ions touch the electrodes and are thus discharged or otherwise disappear from the ion trap. The resonant excitation is usually brought about by an alternating voltage applied across the two end cap electrodes.
The remaining parent ions collect again in a small cloud in the center of the ion trap as a result of the damping in the collision gas. They can now be fragmented. The usual type of fragmentation is collisionally induced decomposition (CID). The relatively soft resonant excitation forces them to oscillate, leading to a large number of low-energy collisions with the collision gas. In many of these collisions, small portions of energy are transferred into the internal structure of the parent ions. The intrinsic energy of the internal molecular oscillation systems increases until one of the weaker bonds within the molecular structure of the parent ion breaks open. A singly charged parent ion forms a daughter ion and a neutral particle; a doubly charged parent ion frequently (but not always) forms two singly charged daughter ions. Since the daughter ions are no longer resonantly excited because they have a different mass and hence a different oscillation frequency, their oscillations are cooled by the collision gas from the moment of cleavage; the daughter ions collect in the center in a small cloud and, according to the present view, do not decompose further. They can then be measured as a daughter ion spectrum in the conventional way by being resonantly and selectively ejected in sequence according to their mass in a detector located outside the ion trap.
Investigations with other types of mass spectrometer have shown that with the harder collision fragmentations used there, not only are two fragment ions created each time, but that these fragment ions can certainly decompose further, presumably in further fragmentation processes or as a result of metastable decomposition, creating granddaughter ions.
We now turn to a field of application in which mass spectrometry plays an important role: proteomics. This frequently involves enzymatic breaking down the proteins to digest peptides, and analyzing the latter by mass spectrometry. If one begins with peptide ions, then so-called internal fragments form in the collision cells; these fragments originate from two cleavages of the chain of amino acids. The incidence of so-called immonium ions here is particularly high; these are charged single amino acids originating from somewhere in the chain. The measurement of such immonium ions has high informational value since they immediately signalize the presence of this amino acid in the peptide. It is frequently possible to read off the amino acid composition of the peptide from the immonium ions, even if it is not possible to thus determine the arrangement of the amino acids along the chain.
It has unfortunately not yet proven possible to measure these immonium ions in ion trap mass spectrometers. If the RF storage voltage used during the fragmentation were low enough for immonium ions to remain in the ion trap after their creation, then the resonant excitation of the parent ions would have to be so weak that they could absorb practically no energy in the collisions; in any case, the cooling effect caused by the collisions is then stronger than the effect of the energy absorption, and no fragmentation occurs. In the case of stronger resonant excitation, the parent ions would then be accelerated as far as the electrodes and they would disappear out of the ion trap.
For fragmentation, the RF storage voltage therefore always has to be quite high, as otherwise there will be no fragmentation. That is a dilemma. The high RF storage voltage produces a high cut-off mass for the storage, and the immonium ions (if they are created at all) cannot be retained. It is usual to choose an RF storage voltage for the fragmentation where the lower cut-off mass for the storage capability is around a third of the mass of the parent ions. It is therefore not only the immonium ions but also smaller ions which are lost from two, three or four amino acids, depending on the size of the peptide.
In a similar way to the immonium ions, other types of light ions produced by fragmentations can also provide information about the structures of the parent ions which is otherwise very difficult to obtain. For example, methods have recently been elucidated which are directed at splitting off ionized derivatization groups (side chains) which have not yet been able to be detected in ion traps using the methods which have been usual until now.